Endocytic Pathways and Intracellular Transport of Aptamer-Drug Conjugates in Live Cells Monitored by Single-Particle Tracking.

Aptamer-drug conjugates (ApDCs) are emerging as targeted therapeutic drugs that can effectively broaden the chemotherapeutic window with higher efficacy and less toxicity. They show promising targeted tumor-killing effects both in vitro and in vivo. However, the mechanisms underlying the cellular internalization and transport of ApDCs remain unclear, and no systematic study on this topic has been reported. Therefore, we herein investigated the endocytic internalization and subsequent transport of ApDCs in mammalian cells through single-particle tracking. We found that ApDC enters the cells mainly by caveolin-mediated endocytosis and that it exhibits cytoskeleton-dependent transport, along microfilaments and microtubules, to acidic endosomes near the cell nucleus in cytoplasm. We also found that the cellular uptake pathways of ApDCs are identical to those of the aptamer itself, confirming that aptamers play a prominent role in the internalization of ApDCs. This study extends our understanding of the internalization and transport process of ApDCs such that the results could serve as the theoretical foundation for designing new ApDCs and, in turn, promoting cancer-targeted therapy.